• 대한생식의학회:학술대회논문집
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• 대한불임학회 2005년도 제49차 추계학술대회
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• pp.123.1-123.1
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• 2005

• Reproductive and Developmental Biology
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• 제35권3호
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• pp.363-367
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• 2011

The purpose of this study was to improve of frozen-thawed sperm using magnetized water in Korean native cattle. Before cryopreservation, without egg-yolk Triladyl$^{(R)}$ solution was flowed though magnet [0, 2000, 4000 and 6000 Gauss(G)] for S min. The freezing of dilluted semen added with Triladyl containing 20% egg-yolk. Analysis of frozen-thawed sperm was estimated viability with SYBR14/PI double stain, membrane intact with hypoosmotic swelling test (HOST), acrosome reaction with FITC-PNA, mitochondria membrane function with Rhodamin 123 by flow- cytometry. Sperm viability was significantly higher in 4000G group than other groups (p<0.05). However, the Hypoosmotic Swelling Test(HOST) was significantly higher in fresh, 4000 and 6000G than 0 and 2000G (p<0.05). In addition, mitochondria membrane damage and acrosome damage were significantly lower in 6000G group than other groups (p<0.05). in conclusion we suggest that magnetized water could be improve ability of sperm on cryopreservation in Korean native cattle.

• Asian-Australasian Journal of Animal Sciences
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• 제20권6호
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• pp.842-849
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• 2007

The purpose of the present study was to produce live offspring from in vitro fertilized goat embryos. Oocytes were collected from abattoir ovaries and kept in oocyte collection medium. Oocytes were washed 4-5 times with maturation medium containing medium-199 with 5 ${\mu}g/ml$ FSH, 100 ${\mu}g/ml$ LH, 1 ${\mu}g/ml$ estradiol-$17{\beta}$ 50 ${\mu}g/ml$ gentamycin, 10% inactivated estrus goat serum, and 3% BSA (fatty acid free). Oocytes were placed in 100 ${\mu}l$ drops of maturation medium containing granulosa cell monolayer and incubated in a 5% $CO_2$ incubator at $38.5^{\circ}C$ for 27 h. For capacitation of spermatozoa fresh semen was processed and mixed in 3 ml fertilization TALP medium containing 50 ${\mu}g/ml$ heparin and kept in the above incubator for 2 h. The capacitated spermatozoa were coincubated with matured oocytes for fertilization. Cleaved embryos were separated and cultured in embryo development medium with oviductal cells and 494 embryos were produced. Recipient goats were synchronized with two injections of 15 mg $PGF_{{2}{\alpha}}$/goat 10 days apart. Eighty early stage embryos were transferred into the uterotubal junction of 14 surrogate mothers using laparoscopy techniques. One recipient delivered twin kids, whereas another two recipients each.delivered a single kid The parentage of these kids was evaluated using highly polymorphic co-dominant microsatellites markers. From the present study, it was concluded that live goat kids can be produced from in vitro matured and fertilized goat embryos, to the best of our knowledge for the first time in India.

• 한국수정란이식학회지
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• 제27권1호
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• pp.21-28
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• 2012

A study was conducted on four crossbred bulls, used as artificial insemination (AI) sires, to correlate their semen quality with their non return rate (NRR). Semen was collected once a week via an artificial vagina, diluted in egg yolk-citrate and maintained at $+7^{\circ}C$ for three days. It was evaluated for sperm motility, viability, morphology immediately after collection and was examined daily for sperm motility, viability and morphology of acrosome, mid piece and tail for a total of three days. A total of 2016 cows were inseminated by two AI technicians. The proportions of sperm with normal heads were 83.4% (63.7~91.7%), the proportion of spermatozoa exhibiting normal morphology (acrosome, mid piece and tail), motility and viability were 89.2% (82.3~92.0%), 71.3% (61.7~75.0%) and 76.7% (65.7~85.0%), respectively in fresh ejaculates. Sperm motility and sperm viability was significantly ($p$ <0.05) lower in Holstein-Friesian ${\times}$ Local bull than in other bulls during all three days of storage. The overall NRR for four bulls was 82.7% (72.9-87.5%). Bulls with higher sperm motility, viability and normal morphology of spermatozoa of individual bull had significantly (each $p$ <0.05) higher NRR. The highest ($p$ <0.01) NRR (87.5%) was observed in a Red Chittagong bull whose semen qualities were significantly ($p$ <0.05) higher than Holstein-Friesian ${\times}$ Local bull (NNR 72.9%). The results of the present study concluded that NRR at 56 days post AI is related to parameters of semen quality. Therefore, semen evaluation may allow the discarding of bulls with poor fertility in an AI program.

• 한국가금학회지
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• 제44권1호
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• pp.59-65
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• 2017

본 연구에서는 닭 정액의 운동성 분석에 미치는 요인을 분석하기 위하여 희석배율에 따른 생존성을 조사하고, 신선정액의 희석 시에 필요한 조건을 확보하기 위하여 희석제에 단백질원으로서 BSA와 FAF-BSA의 첨가가 닭정자의 생존성에 미치는 영향을 관찰하였다. 최종 희석율이 1/4이 되도록 닭 정액을 희석하여 생존성을 조사하면 신선정액에서 희석율이 1/8, 1/16 및 1/32로 낮아질수록 89.9%, 69.9% 및 53.2%로 생존율이 낮아지는 것을 관찰하였으며, 단백질원으로 BSA나 FAF-BSA를 이용하면 생존율을 유의적으로 증진시킬 수 있음을 증명하였다. 또한 이러한 현상은 동결정액의 희석에도 중요한 요인으로 관찰되었는데, FAF-BSA를 이용하여 희석할 때 단백질이 없는 경우보다 양호한 생존율이 나타난다는 것을 관찰하였다(17.6% 대 34.0%). 그러나 단백질원을 활용하더라도 닭정액의 희석율이 낮아질수록 생존율이 저하하는 현상은 지속적으로 나타나고 있으며, 신선정액에 비하여 동결정액은 장수성이 낮은 것으로 추정되었다. 그러므로 본 연구에서 사용된 정자의 분석방법은 닭 정액동결을 실시하여 운동성을 분석할 때 필요한 기본 정보를 제공할 수 있을 것으로 판단되며, 닭 희석배율을 낮추어 동결정액의 효율성을 높이는 연구에 기여할 것으로 추정된다.

• 한국수정란이식학회지
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• 제16권2호
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• pp.139-144
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• 2001

본 연구는 숫소 개체별, 정자의 형태, 정자의 전처리 및 난자의 투명대부착과 미부착별로 현미조작에 의해 난자의 세포질내 단일정자를 주입했을 때 웅성전핵 형성율과 체외발생율을 조사하였다. 1. 숫소 개체별 정자를 이용하여 ICSI를 하였을 때 웅성전핵 형성율은 각각 73.9∼87.0%였고 체외발생율은 33.3∼60.9%를 나타냈다. 2. 신선 및 동결정자와 미부절단 정자와 두부, 미부손상 정자를 이용하여 ICSI를 하였을 때 웅성전핵 형성율은 각각 82.0%, 78.0% 42.2%, 51.1%였고, 체외발생율은 56.0%, 42.0%, 17.8%, 22.2%로서 신선정자를 이용했을 때가 다른 처리군에 비해 높은 전핵형성율과 체외발생율을 나타냈다. 3. 정자를 heparin, BFF, His, Ca Ionophore, I + caffeine 액으로 처리후 ICSI를 하였을 때 전핵 형성율은 66.7∼82.2%였고, 체외발생율은 33.3∼60.6%로서 I + caffeine 처리법이 다른 처리군에 비해 높게 나타났다. 4. 투명대 부착 난자 및 미부착 난자로 ICSI를 하였을 때 전핵형성율은 각각 80.0%, 72.0%였고, 체외발생율은 46.0%, 36.0%로서, 토명대부착 난자가 투명대 미부착 난자에 비해 높은 웅성전핵 형성율과 체외발생율을 나타냈다.

• 한국수정란이식학회지
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• 제27권1호
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• pp.29-35
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• 2012

The purpose of this study was to examine the effect of growing stages of the Korean Native Striped Bull (KNSB) on the freezability and fertility of frozen-thawed semen. First, we investigated the total motility (TM) and progressive motility (PM) according to the diluent used for semen freezing. Second, we examined the effect of the age of KNSB on semen volume, TM and PM of fresh and frozen-thawed semen. Third, we examined the effect of frozen semen from the different age of KNSB on the $in-vitro$ fertilization rate, and the artificial insemination pregnancy rate. The diluents used in this experiment were Triladyl$^{(R)}$ and Tris-egg yolk extender (EYE). Semen was collected from 5 KNSB in the growing stage (15 months) and 5 adult KNSB (36 months). When Triladyl or Tris-EYE extender was used for semen freezing, there was no difference of the mean TM and the mean PM. However, the mean TM was significantly higher in Bull No. 1885 than Bull No. 4283 ($p$ <0.05). The mean volume of semen collected from the 15-month-old bulls (2.3 ml) was significantly lower ($p$ <0.05) than that from the 36-month-old bulls (5.0 ml). The mean semen concentration was similar for the 15-month-old ($2.1{\times}10^9$ spermatozoa/ml) and 36-month-old ($1.8{\times}10^9$ spermatozoa/ml) bulls. For the 15-month-old and 36-month-old bulls, the mean TM of fresh semen were 93.7% and 88.3%, respectively, and the mean PM were 97.0% and 88.3%, respectively; the 15-month-old bulls showed a particularly high PM ($p$ <0.05). For the 15-month-old and 36-month-old bulls, the mean TM (56.0% and 58.0%, respectively) and the mean PM (64.0% and 70.7%, respectively) of frozen-thawed semen did not differ. The development rates of embryos after $in-vitro$ fertilization and the pregnancy rate after artificial insemination using frozen-thawed semen did not differ according to the bull's age. In summary, semen volume differed according to the bull's age, but semen concentration and survival rate, the $in-vitro$ fertilization rate, and the pregnancy rate did not differ according to the stripe bull's age. Accordingly, semen from bulls in the growing stage can be collected and frozen for the preservation and multiplication of rare livestock.

• 한국가금학회지
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• 제48권4호
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• pp.185-191
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• 2021

닭 정자는 저온 환경에서 생존하는 능력을 보유하고 있으나 정자의 운동성과 첨체 손상에 대한 연구는 상세히 알려져 있지 않다고 판단된다. 본 연구에서는 닭 정자를 BPSE 및 Lake 희석액으로 희석하고 보존 용기를 1.5 mL e-tube와 0.5 mL 스트로에 보존하여 정자 장수성을 조사하였다. 밀봉된 0.5 mL 스트로에 보존된 정자의 운동성이 3일동안 저온보존 시 더 높은 것으로 관찰되었다(68.6±3.1% vs. 22.1±5.7%). BPSE로 희석하여 0.5 mL 스트로에 보존된 닭 정자의 운동성 또한 Lake 희석액보다 더 높게 관찰되었다(57.7±5.6% vs. 37.7±5.4%). 첨체 온전성도 0.5 mL 스트로에 보존하는 것이 우수하였고 냉장보존일이 증가함에 따라 손상된 첨체 비율이 증가하였다. 본 연구에서 0.5 mL에 밀봉된 닭 정자는 저온 보존에 이용될 수 있으며 정자의 운동성을 높여주며 첨체 온전성을 증가시키는 것으로 관찰되었다.

• 한국수정란이식학회지
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• 제17권1호
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• pp.55-59
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• 2002

ICSI시 동결 융해한 부고환 정자의 이용 가능성을 알아보고자 난자의 배양시 체외성숙율과 활성화 처리를 한 난자와 동결 융해한 부고환 정자로 ICSI시 체외발생율을 조사하였으며, 결과를 요약하면 다음과 같다. 1. 난포란을 회수 후 24시간 배양하였을 때 배양 시간에 따른 GV, MI, M II로의 체외성숙율은 각각 7/60(11.7%), 5/60(8.3%), 48/60(80.0%)였고 30시간 배양 시간에 따른 GV, MI, M II로의 체외성숙율은 각각 3/60(5.0%), 4/60(6.7%), 53/60(88.3%)였고 퇴화란은 각각 2/60(3.3%)와 1/60(1.7%)였다. 2. 동결 융해한 부고환 정자를 이용하여 활성화 처리를 한 난자에 ICSI를 하였을 때 상실배와 배반포로의 체외발생율은 각각 12/46(26.1%), 22/46 (47.8%)로서 비활성화처리 난자군 5/39 (12.8%), 10/39(25.6%)에 비해 높은 체외발생율을 나타냈다. 3. 활성화 처리를 한 난자에 신선정자, 부고환 정자 및 동결 융해한 부고환 정자로 ICSI시 체외 발생율은 각각 24/45(53.3%), 15/40(37.50%), l1/43 (25.6%)로서 신선정자에 비해 동결 융해한 부고환 정자처리군은 체외발생율은 약간 낮았지만 이용 가능성이 있음을 확인하였다.

• Clinical and Experimental Reproductive Medicine
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• 제21권1호
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• pp.1-11
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• 1994

The occurrence and time course of capacitation, acrosomal loss, and hyperactivated motility require quantitative definition in order to characterize fertile human sperm. Recently the method has been developed to estimate the quality of spermatozoa by using kinematic parameters such as curvilinear velocity(VCL), average path velocity(VAP), linearity(LIN), straightness(STR), amplitude of lateral head displacement(ALH), and beat cross frequence(BCF) from Computer Assisted Sperm Analysis (CASA). In this study, using the Hamilton Thorn motility analyzer HTM 2030(Hamilton Thorn Research, Beverly, MA), we attempted to identify the spermatozoa with hyperactivated motility (HA) objectively and to monitor hyperactivation of human spermatozoa during incubation in capacitating media and after treatment of calcium ionophore as compared with acrosome status. And we examined whether HA are related to the result of SPA. Semen samples obtained from 16 healthy men were prepared by swim up technique and preincubated in a capacitating media(modified BWW medium) for 5 hours and treated with calcium ionophore solution. The acrosome reaction was detected with PSA-FITC labelling of the acrosome and in vitro sperm ferilizing capacity was assessed by the zona free hamster ovum penetration assay (SPA). The incidence of hyperactivated sperm was 2.6% in fresh semen, 14.3% of the swim up population, 13.7% after 5h of incubation. Significant increase of percentage of hyperactivated sperm was observed after the incubation (p<0.05) but after treatment, no significant changes of percentage of hyperactivated sperm(l1.8%) in contrast to significant rise in the percentage of acrosome reacted cells. Correlation analysis failed to show any significant relationship between the percentage of sperm with HA and SPA score. In conclusion, although no direct correlations were found between the results of SPA and HA, hyperactivation of sperm is associated with capacitation and monitoring hyperactivated sperm will be expected as a method of evaluating the functional quality of sperm such as SPA.